RU2769129C1 - Fermentation plant for cultivation of methylococcus capsulatus methane-oxidizing bacteria - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: invention relates to the field of biotechnology. The fermentation plant for M.capsulatus bacteria includes a fermenter for aerobic cultivation processes, equipped with gas and liquid process flow supply pipes, outlet pipes for the obtained biomass, waste gas and gas-liquid mixture, elements for organizing flows inside the fermenter, additionally contains a storage tank connected to the fermenter by an equalizing gas line in its upper part, a gas separator, an ejector and a gas-liquid flow velocity equalization line, in addition, the elements for organizing flows inside the fermenter are located in the lower part of the fermenter and are made in the form of two sparge rings connected to the natural gas and oxygen supply lines and a pressure inlet pipe of the gas-liquid mixture connected to the ejector.

EFFECT: invention makes it possible to increase the reliability and efficiency of the device by eliminating accidents during fermentation and increasing the use of natural gas entering the fermenter in the process of biomass synthesis.

1 cl, 1 dwg, 1 ex.

Description

The invention relates to the field of biotechnology, in particular to devices for cultivating the biomass of methane-oxidizing microorganisms Methylococcus capsulatus.

The relevance of developing a new efficient bioreactor in relation to gas-liquid fermentation on natural gas of aerobic methane-oxidizing microorganisms is associated, on the one hand, with an increased economic interest in the use of significant reserves of natural gas, and on the other hand, with inefficient instrumentation of the main stage of protein production - the fermentation process.

The use of natural gas will make it possible to create biotechnological installations that provide agro-industrial complexes and fish farming with complete protein feed, as well as the production of a wide range of bioproducts and their further processing.

Despite a significant number of domestic and foreign hardware solutions, the specificity of the process using two sparingly soluble gases, oxygen and methane, has not made it possible to solve the problem of fermentation and protein production from natural gas in industrial bioreactors sufficiently efficiently and energy-efficiently. (Vinarov A.Yu. et al. “Fermentation apparatus for microbiological synthesis processes”, M., DeLi Print, 2005, 275 p.).

There is a wide variety of designs of bioreactors for aerobic cultivation using traditional mixers, ejectors, circulation pumps and bubblers that can be used to grow the biomass of metautilizing microorganisms, however, in most cases, their design characteristics and energy-consuming indicators make this process inefficient, which determines the task of developing a new apparatus for aerobic cultivation of metautilizing microorganisms is relevant and practically important.

As a result of the patent information search, the following patents were selected.

Known (GB1353008A, class C12M1/04; C12M1/08, publ. 1974-05-15) apparatus for obtaining biomass of microorganisms, which uses the principle of aeration and mixing of the fermentation medium. The known apparatus is made of two vertically arranged containers, between which the circulation of the entire fermentation medium is organized due to the difference in the densities of the aerated and degassed liquid. Aeration is carried out using a bubbler located in the lower part of one of the tanks, and the exhaust gas separated from the liquid as a result of its degassing exits through the upper pipe on another vertical tank, through which the degassed liquid circulates down, passing the heat exchange device. The device is designed to work with large volumes of fermentation medium.

The disadvantages of this apparatus include the insufficient degree of dispersion of the gas phase and the turbulence of the fermentation medium, which does not allow the effective use of poorly soluble gaseous substrates and dispersed media for fermentation processes, as well as the relatively low contact surface of the gas-liquid phases, due to the coalescence of rising bubbles, due to than, in this apparatus, the transport of gaseous sources of cell growth does not ensure their effective development and increase in cell biomass. In addition, the presence of the problem of difficult solubility of gases leads to their excessive consumption and possible disruption of the fermentation process, due to the fact that the gas-liquid mixture obtained as a result of the fermentation reaction can escape from the apparatus, leading to its shutdown or emergency situations.

Known apparatus for growing microorganisms (RU 2352626, class C12M 1/02, C12M 1/04, publ. 20.04.2009). The device contains a container filled with culture liquid to a certain level and equipped with nozzles for supplying liquid mineral nutrient medium, air and removing accumulated biomass, as well as at least one additional container, which is an absorber of a gaseous substrate. These containers have flared shells installed along the axis of the container, which serve to separate the liquid-filled part of the container into lifting and lowering channels. Bubblers for air supply are located in the lifting channels of the tanks. The tanks are connected in the lower part by a liquid pipeline in such a way that the downstream channel of the main tank is connected to the lifting channel of the additional tank, and the downstream channel of the additional tank is connected to the upstream channel of the main tank, and a liquid circulation booster is installed in the liquid line between the tanks. The flanging in the upper part of the shell of the additional container can be made in the form of a funnel directed inside the downcomer, symmetrical to the axis of the downcomer. A breathing pipe can be installed along the center of the flanging, connecting the supra-liquid part of the container with the inside of the downcomer, and another funnel is installed below this funnel and the breathing pipe, coaxially with them. EFFECT: invention makes it possible to obtain a protein mass of aerobic microorganisms (gaprin) when using a gaseous substrate (natural gas) as a food for microorganisms.

The main disadvantages of this apparatus are the high energy costs for the circulation of a large amount of liquid between the tanks; low degree of dispersion of sparingly soluble gases of air (oxygen), methane in the aquatic environment (with a bubble size of about 10 mm), achieved in gravity-type ejectors; the complexity of the design of the fermenter; the low rate of mass transfer of sparingly soluble gases and the associated low productivity in terms of biomass do not allow efficient use of this apparatus in industry for obtaining biomass of methane-assimilating bacteria. Also, the disadvantages include the difficulty in adjusting the gas outlet from the apparatus due to the formation of a large amount of gas-liquid mixture, which creates excess pressure inside the apparatus, leading to the “flooding” of the fermenter, requiring its emergency shutdown.

Known fermentation unit for methane-oxidizing bacteria (RU 2580646, class C12M 1/02, C12M 1/04, C12M 1/36, C12Q 3/00, publ. 10.04.2016), selected as a prototype. The fermentation plant for methane-assimilating microorganisms is designed to operate at an overpressure of 0.1-1.0 MPa and contains a column fermenter for carrying out aerobic cultivation processes and two reactors connected to it by pipelines of gas and liquid process streams. The fermenter contains a housing, in the upper part of which there is a branch pipe for the outlet of the exhaust gas and a branch pipe for supplying nutrient salts and process water, nutrition components and seed biosuspension. In the lower part, there is a branch pipe for inlet of the gas-liquid flow into the fermenter from the reactors and a pipe for the biosuspension output from the fermenter to the reactors. Each reactor contains a branch pipe for extracting a gas-liquid flow from the fermenter, installed below the level of the culture liquid in the reactor, a disk mixer with a drive, installed at the level of the culture liquid in the reactor, to organize flows inside the fermenter, a branch pipe for supplying biosuspension from the fermenter to the reactor. The first reactor contains a methane-containing gas supply pipe connected in the upper part. The second reactor contains an oxygen-containing gas supply pipe connected in the upper part. The outlet branch pipe from the exhaust gas column fermenter is made with the possibility of connection to a defoaming device. The branch pipe is installed with the possibility of connection to the line for transferring the exhaust gas for combustion or for re-supply of gas to the reactors.

The disadvantages of the known installation include both structural complexity and complexity in operation. In addition, in this fermenter, the complete separation of the gas from the liquid phases during the fermentation of the finely dispersed state of the gas phase in the volume of the fermenter is insufficient. Regulation of gas and liquid flows is carried out inefficiently, which leads to an excess of gas consumption, as well as to unauthorized shutdowns of the apparatus and emergency situations. Putting the fermenter into operation is a long and laborious process.

The problem of the invention is the development of a fermentation plant for the cultivation of methane-oxidizing bacteria Methylococcus capsulatus with the possibility of stabilizing the start-up modes of the fermentation plant while reducing the sequence of operations, due to the most complete separation of the gas phase from the liquid, eliminating excessive pressure from the fermenter of the gas-liquid mixture and its "flooding with liquid".

The technical result of the invention is to increase the reliability and efficiency of the device by eliminating emergency situations in the fermentation process and increasing the degree of use of natural gas entering the fermenter in the process of biomass synthesis.

The problem posed and the technical result are achieved by the fact that the fermentation plant for methane-oxidizing bacteria Methylococcus capsulatus includes a fermenter for carrying out aerobic cultivation processes, equipped with nozzles for supplying gas and liquid process streams, outlet pipes for the resulting biomass, exhaust gas and gas-liquid mixture, elements for organizing flows inside the fermenter . According to the invention , the installation further comprises a storage tank for removing the resulting biomass from the fermenter, a gas separator, an ejector and a gas-liquid flow rate equalization line, including gas flow lines with control valves. One gas flow line connects the top of the fermenter to the ejector and the other connects the top of the fermenter to the exhaust gas outlet. The fermenter and the gas separator are connected by a line for the output of the gas-liquid mixture from the fermenter. The ejector and the gas separator are connected to each other from above along the gas flow line with a control valve, and from below - along the liquid flow line through a heat exchanger and a centrifugal pump. Elements for organizing flows inside the fermenter are located in the lower part of the fermenter and are made in the form of two bubbling rings connected to natural gas and oxygen supply lines and a gas-liquid mixture pressurized inlet pipe connected to the ejector.

Introduction to the installation of a line for equalizing the speeds of the gas-liquid flow, including a gas flow line with a control valve connecting the fermenter with the exhaust gas outlet pipe, makes it possible to ensure the stability of the fermenter, which has a positive effect on the efficiency of the fermentation process, as well as an increase in the degree of utilization of natural gas, to eliminate failure in installation operation.

Execution of elements for organizing flows inside the fermenter in the form of two bubbling rings connected to natural gas and oxygen supply lines and a gas-liquid mixture pressurized inlet connected to the ejector ensure the rotation of the flow inside the fermenter, creating optimal conditions for the fermentation process, thereby increasing efficiency biomass synthesis.

The device is illustrated in the drawing, which shows the installation for methane-oxidizing bacteria Methylococcus capsulatus.

The fermentation plant includes a fermenter 2 for growing bacteria, a gas separator 2, and a storage tank 3. In the lower part of the fermenter 1 there are bubbling rings 4 and 5 connected to lines 6 and 7 for supplying natural gas and oxygen, respectively. The gas separator 2 is connected by a line 8 through a heat exchanger 9 and centrifugal pump 10 with an ejector 11 of the liquid-gas type. The line for equalizing the speeds of the gas-liquid flow, including the gas flow line 12 with the control valve 13, connecting the upper part of the fermenter 1 with the ejector 11 and the line 14 with the control valve 15, connecting the upper part of the fermenter 1 with the branch pipe 16 for exhaust gas outlet, equipped with a pressure regulator 17 . Fermenter 1 pipe 18 - outlet of the obtained biomass is connected to the storage tank 3, equipped with a pipe 19 outlet of the resulting biomass. The heat exchanger 9 includes an inlet 20 of cooling water and an outlet 21 of water from the heat exchanger 9. A pipe 22 for supplying process water, a pipe 23 for supplying an aqueous ammonia solution to the fermenter, a pipe 24 for supplying a solution of nutrient salts and a pipe 25 for the outlet of a gas-liquid mixture are connected to the body of the fermenter 1 . The gas separator 2 is connected from above by a line 26 for the outlet of a part of the exhaust gas (exhaust gas), equipped with a control valve 27 - with an ejector 11. From the ejector 11 through the pipe 28, the gas-liquid mixture under pressure enters the lower part of the fermenter 1.

Fermentation plant for the cultivation of methane-oxidizing bacteria Methylococcus capsulatus works as follows.

Natural gas and oxygen enter the fermenter 1 through bubbling rings 4 and 5 connected to pipes 6 and 7. To ensure the growth of biomass, a working solution of nutrient salts is supplied to the fermenter 1 through the pipe 24. To maintain the pH in the fermenter 1, an aqueous solution of ammonia is fed through the pipe 23. Also, process water is supplied to the fermenter 1 through the pipe 22 to organize the flow in the fermenter. The gas-liquid mixture from the lower part of the fermenter 1 through the pipe 25 due to the pressure difference enters the gas separator 2. From there, part of the exhaust gas (off-gas) is removed from the process through the pipe 16, and the remaining amount of gas through line 26 enters the ejector 11 and is introduced back into the fermenter 1 The pressure in the gas separator 2 is maintained by a pressure regulator 17. The liquid part of the flow from the gas separator 2 exits through the branch pipe of the line 8, passes through the heat exchanger 9 to maintain the required liquid temperature, and with the help of a centrifugal pump 10 enters the ejector 11 under pressure. From the gas separator 2 along the line 26 a controlled amount of the gas mixture is output and enters the ejector 11. The liquid is constantly supplied there by the pump 10 from the gas separator 2 through the pipe along the line 8. In the ejector 11, a gas-liquid mixture is formed with a given distribution of gas bubbles in the liquid. And through the nozzle 28, the resulting mixture enters the fermenter 1 under pressure, ensuring the rotation of the flow inside it. With the help of the control valve 15, the speeds of the gas-liquid flow from the fermenter 1 to the gas separator 2 and the liquid flow from the gas separator 2 to the heat exchanger 9, pump 10 and ejector 11 are equalized.

Equalization of the rates of movement of the gas-liquid flow from the fermenter 1 to the gas separator 2 and the liquid flow from the gas separator 2 to the heat exchanger 9 is carried out due to the control valve 15. When the valve 15 is closed, the gas flow from the fermenter 1 from the pipe 14 to the gas separator 2 decreases, which also contributes to an increase flow of the gas-liquid mixture from the fermenter 1 through the nozzle 25 into the gas separator 2 and, as a result, the level of the working liquid medium in the gas separator 2 rises. mixture from the fermenter 1 through the nozzle 25 to the gas separator 2 and, as a result, increase the level of the working liquid medium in the gas separator 2. Setting the required position of the valve 15 provides the required level in the gas separator 2, and as a result, the optimal mode of gas separation in it.

The optimal ratio of gas flows, ensuring an increase in the degree of use of natural gas, is also created by regulating the degree of opening of the valve 15 installed on the line 14 connected with the branch pipe 16 with the outlet of the exhaust gas, and by regulating the valves 13 and 27 on the outlet lines of the part of the exhaust gas (off-gas) into the ejector 11. The control valves 13, 15, 27 and the pressure regulator 17, installed on the gas-liquid flow rate equalization line, ensure reliable trouble-free operation of the entire installation, excluding "flooding" of the gas-liquid mixture of the gas separator 2 and prevent emergency situations and unplanned shutdowns that require further commissioning work.

The resulting biomass from the fermenter 1 enters through the nozzle 18 into the storage tank 3, where it is accumulated and removed from the fermentation system through the nozzle 19. The pressure in the storage tank 3 and the fermenter 1 is equalized by connecting them with an equalizing gas line in the upper part of the fermenter 1 and storage tank 3.

The principal advantage of the given fermentation plant for cultivating methane-oxidizing microorganisms in comparison with the previously known ones is the most complete separation of the gas phase from the liquid in the gas separator 2 by means of forced intake by its ejector 11 of the liquid-gas type. As a result, there is a decrease in the head loss of the centrifugal pump 10 due to gas saturation, thereby increasing the efficiency of the ejector 11, which increases the degree of saturation of the liquid introduced into the fermenter 1 with the returned gas mixture (off-gas).

An example of the technological mode of cultivation

The culture of Methylococcus capsulatus is grown in the fermenter 1. The cultivation process is carried out in a continuous mode at excess pressure. To carry out the cultivation process, the fermenter 1 is filled with process water and a solution of mineral nutrient salts. Natural gas with a methane content of at least 90% is used as the main technological raw material for the growing process. The process is carried out in an environment saturated with oxygen. As a source of oxygen for supply to the fermenter 1, both pure gas and oxygen-enriched air can be used.

In the fermenter 1 maintain the pH of the environment, set in the range of 5.6-5.8 units. This pH range in the apparatus is maintained by supplying ammonia water to the system, which is also a source of nitrogen. The operating temperature of the process is maintained in the range of 42-45°C by cooling the circulating biomass in an external heat exchanger 9. The operating pressure in the apparatus is 0.5 MPa.

The average working concentration of cells in the selected biomass is 18÷21 g/l (ASV), with a specific flow rate of the medium through the apparatus is not less than 0.2 h ^-1 .

In the gas separator 2, at least 95% (by volume) of the gas phase is separated from the gas-liquid mixture, which makes it possible to maintain the stability of the operation of the centrifugal pump 10, i.e. to ensure the preservation of its main characteristics - productivity and pressure.

The organization of forced gas extraction from the gas separator 2 using the ejector 11 and its introduction as part of the gas-liquid mixture into the fermenter 1 makes it possible to ensure a high degree of dispersion of the gas phase in the liquid inside the apparatus and increase the degree of use of natural gas.

Claims (1)

Fermentation plant for methane-oxidizing bacteria Methylococcus capsulatus, including a fermenter for carrying out aerobic cultivation processes, equipped with nozzles for supplying gas and liquid process streams, outlet pipes for the obtained biomass, exhaust gas and gas-liquid mixture, elements for organizing flows inside the fermenter, characterized in that the installation additionally contains a storage tank for removing the resulting biomass from the fermenter, connected to the fermenter by an equalizing gas line in its upper part, a gas separator, an ejector and a gas-liquid flow rate equalization line, including gas flow lines with control valves, one of which connects the upper part of the fermenter with the ejector, and the other is the upper part of the fermenter with a branch pipe for the outlet of exhaust gas, and the fermenter and the gas separator are connected by a line for the output of the gas-liquid mixture from the fermenter, the ejector and the gas separator are interconnected at the top along gas flow lines with a control valve, and from below - along the liquid flow line through a heat exchanger and a centrifugal pump, in addition, the elements for organizing flows inside the fermenter are located in the lower part of the fermenter and are made in the form of two bubbling rings connected to natural gas and oxygen supply lines and a branch pipe for inlet under pressure of a gas-liquid mixture associated with the ejector.

Images